Summary: Dendritic cells (DC) are classically regarded as "professional" antigen presenting cells because they have immense capacity for capturing and processing protein antigens. They subsequently display peptide fragments of the antigen on their surface in association with MHC-I/II molecules and present this complex to MHC compatible T cells. Although they lack phagocytic function, they are powerful activators of B and T lymphocytes and provide essential accessory functions for T cell mediated cytotoxicity. These cells have become popular cells in the field of tumor vaccines. However, as their use increases, a number of scientific issues need to be addressed. These include: 1. Identity: What surface markers identify dendritic cells? What would be an acceptable test of identity for these cells to ensure lot-to-lot consistency if multiple administrations are planned? 2. Functional Assays: How would potency for activated DC be evaluated? Would an mixed lymphocyte reaction or generation of activated antigen specific responder cells be sufficient? 3. How we can determine if dendritic cells are really pulsed? Should attempts be made to quantitate the petide load or whether peptide is presented in the context of MHC? Our new project will address some or all of these questions. Our research activity this period included optimization of generation of human dendritic cells from elutriated monocytes obtained from normal donors at the NIH Blood Bank. We compared the phenotype of dendritic cells activated by GM-CSF with either IL-4, IL-13 or IL-13 agonist. In most cases, we were able to generate dendritic cells, however, these cells did not seem to express CD83 antigen, an important marker for maturation. We also tested effect of various culture conditions on these cells. Currently, it is planned to test various growth medias and use of other cytokine mixtures for optimal maturation of these cells. Attempts are also being made to identify expression of new antigens or receptors on dendritic cells.